Abstract
Here we report fabrication of Gelatin-based biocomposite films and their application in developing epithelial patches. The films were loaded with an epithelial cell growth factor cocktail and used as an extracellular matrix mimic for in vitro regeneration of organized respiratory epithelium using Calu-3 cell line and mesenchymal stem cells (MSCs). Our data show differentiation of Calu-3 cells on composite films as evidenced by tight junction protein expression and barrier formation. The films also supported attachment, migration, and proliferation of alveolar basal epithelial cell line A549. We also show the suitability of the composite films as a biomimetic scaffold and growth factor delivery platform for differentiation of human MSCs to epithelial cells. MSCs differentiation to the epithelial lineage was confirmed by staining for epithelial and stem cell specific markers. Our data show that the MSCs acquire the epithelial characteristics after 2 weeks with significant reduction in vimentin, increase in pan cytokeratin expression, and morphological changes. However, despite the expression of epithelial lineage markers, these cells did not form fully functional tight junctions as evidenced by low expression of junctional protein ZO1. Further optimisation of culture conditions and growth factor cocktail is required to enhance tight junction formation in MSCs-derived epithelial cells on the composite hydrogels. Nevertheless, our data clearly highlight the possibility of using MSCs in epithelial tissue engineering and the applicability of the composite hydrogels as transferrable extracellular matrix mimics and delivery platforms with potential applications in regenerative medicine and in vitro modelling of barrier tissues.
Highlights
Respiratory tissue injuries, infections and degeneration lead to several serious pathologies such as Chronic Obstructive Pulmonary Disease, Asthma, and Chronic Bronchitis
The hyaluronic acid (HA) is an important component of extra cellular matrix in various tissues in human body, such as connective, epithelial, and neural tissues Due to its high water retention capacity, the presence of HA provides the advantage of increased volume for the substrate which substantially increase its ability to be loaded with growth factors
In a previous work from our lab (Knopf-Marques et al, 2017), we have demonstrated that the stability of Gelatin-Hyaluronic Acid-Tyramine films (Gelatin)/HA membrane films can be improved using HA derivative such as HA-tyramine by creating an interpenetrated network through a double crosslinking step
Summary
Respiratory tissue injuries, infections and degeneration lead to several serious pathologies such as Chronic Obstructive Pulmonary Disease, Asthma, and Chronic Bronchitis. The extracellular matrix (ECM)-based scaffolds are potential tools for in vitro tissue development due to their ability to mimic the native microenvironment, e.g. collagen type I, collagen IV, laminin, and glycoproteins (Yen, Chan, & Lin, 2010) These ECM components induce epithelial migration via integrin signalling and are through to play a key role in directing epithelial repair. The airway epithelial regeneration in the presence of thin biomaterial substrates is considered as one of the possible methods to induce airway epithelium formation for developing robust models which have the basement membrane component (Vrana et al, 2011) In this context, Gelatin based biomaterials have been used in supporting the growth of liver(Yan et al, 2005), bone (Yang, Hsu, Wang, Hou, & Lin, 2005), cardiac (Pok, Myers, Madihally, & Jacot, 2013) and skin (Lu, Oh, Kawazoe, Yamagishi, & Chen, 2012) tissues. The system was used to assess whether it is capable to support the differentiation of the BM-MSCs towards respiratory epithelium lineage
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